Alcohol abuse contributes significantly to disease burden and mortality in the United States, with an annual healthcare cost of $223 billion and death toll of nearly 88,000. It is well known that chronic alcohol consumption leads to gut injury, malnutrition/malabsorption, and increased incidence of colorectal cancers. Studies have implicated quantitative and qualitative alterations of gut microbiota as well as dysregulation of tight junction proteins as key contributors to gut injury, but our understanding of the mechanisms underlying ethanol-induced pathologies remains incomplete. Gut homeostasis is achieved through tightly regulated interactions between epithelial cells, immune cells, and the microbiome. Deeper understanding of the interplay among these three components is necessary to fully identify processes of alcohol-induced gut injury. However, studies to date have not fully uncovered ethanol-induced changes in epithelial cell gene expression or specific microbial shifts that are responsible for tissue injury. More importantly, dose- and region- specific changes have been largely understudied despite the fact that they are likely to play distinct roles in the initiation or exacerbation of various aspects of ethanol-induced pathophysiology. In this application, we will simultaneously define ethanol-mediated changes in the (i) gut microbiome, (ii) mucosal and immune cell gene expression, and (iii) barrier function within all major sections of the gut (duodenum, jejunum, ileum, and colon) using a translational nonhuman primate model of voluntary ethanol self-administration. This unique animal model provides us with an unprecedented opportunity to investigate region- and dose-dependent alterations in gut homeostasis caused by alcohol. Completion of these studies will provide us with the knowledge necessary to design interventions to repair alcohol-induced tissue injury.